The present invention relates to a method of depleting normal and transformed lineage committed cells from a sample from a patient with chronic myeloid leukemia.
Chronic myeloid leukemia (CML) is a monoclonal expansion of a transformed pluripotent stem cell (Fialkow et al., 63:125, 1977, American Journal of Medicine). Myeloid cells, erythroid cells and less frequently lymphocytes arise from the leukemic clone (Bakhshi et al., New Eng. J. Med. 309:826, 1983). CML is characterized in more than 90% of patients by the rearrangement between the break cluster region (BCR gene, located on chromosome 22) and the ABL gene (located on chromosome 9) (Bartran et al., Nature 306:277, 1983).
Although patients with CML may have a prolonged course, the disease is invariably lethal. Bone marrow transplantation is the treatment of choice for this patient population with a curative rate of 90% in some centres. However, for 60% of patients this therapy may not be available either due to the lack of a suitable donor due to differences in human leukocyte antigens (HLA) or the age of the recipient.
For these reasons, other treatment options have been evaluated for their ability to remove the leukemic cells from the harvested material without depleting or damaging the co-existing benign (non-malignant) stem cells. The methods have included various drug regiments (Degliantoni et al., Blood 65:753, 1985) or the culture of the patient cells using the Dexter (long term culture) system which was shown to preferentially support the proliferation of benign stem cells as compared to malignant cells (Barnett et al., Bone Marrow Transplant 4:345, 1985).
Clinical experience has confirmed that although the leukemic burden has been greatly reduced using such protocols, the malignant cells in most patients have not been entirely eradicated and patients relapse with their original disease. (Coutintro et al., Progress in Clinical and Biological Research 333:415, 1990 and Deisseroth et al., Blood 83:3068, 1994) In addition, the high incidence of graft failure also suggests that certain types of treatment may have had adverse effects on the non-malignant stem cells (Talpaz et al., Blood 85:3257, 1995 and Daley and Goldman, Exp. Hematol. 21:731, 1993).
Further analysis of this disease has focussed on dissecting out certain populations of primitive cells in an attempt to understand at what stage the clonal abnormality occurs (Verfaille et al., Blood 87:4770, 1996). These studies may be limited by the low frequency of primitive cells due to the clonal proliferation of lineage committed cells. Further studies of this disease may be facilitated if the mature lineage committed xe2x80x9ccontaminatingxe2x80x9d cells could be reduced or eliminated.
The present inventors have developed an antibody composition for use in preparing cell preparations depleted of normal and transformed lineage committed cells, for example from blood or bone marrow samples from patients with chronic myeloid leukemia. The antibodies in the antibody composition are specific for selected markers associated with lineage committed cells. In particular, the present inventors have found that using an antibody composition containing antibodies specific for glycophorin A, CD2, CD3, CD14, CD15, CD16, CD19, CD24, CD56, CD66b and IgE gives a cell preparation enriched for hematopoietic stem cells and progenitor cells and depleted of committed lineage or differentiated cells.
Accordingly, the present invention relates to an antibody composition comprising antibodies specific for glycophorin A, CD2, CD3, CD14, CD15, CD16, CD19, CD24, CD56, CD66b and IgE which gives a cell preparation depleted of lineage committed cells.
The present invention also provides an antibody composition comprising antibodies specific for glycophorin A, CD2, CD3, CD14, CD16, CD19, CD24, CD56, CD66b and IgE. Such a composition can be used in combination with antibodies to CD15 to prepare a cell preparation depleted of lineage committed cells.
The present invention also includes a negative selection method for depleting lineage committed cells from a sample from a patient with chronic myeloid leukemia comprising:
(a) reacting the sample with an antibody composition comprising antibodies specific for glycophorin A, CD2, CD3, CD14, CD15, CD16, CD19, CD24, CD56, CD66b and IgE under conditions so that conjugates between the antibodies and cells in the sample having the antigens glycophorin A, CD2, CD3, CD14, CD15, CD16, CD19, CD24, CD56, CD66b and IgE on their surfaces;
(b) removing the conjugates; and
(c) recovering a cell preparation which is depleted of lineage committed cells.
The antibody composition of the invention may be used to prepare cell preparations from patients with chronic myeloid leukemia that are depleted of matured differentiated or lineage committed cells and can withstand freezing.
In a preferred embodiment, the sample is first treated with an antibody to CD15 and then it is treated with a cocktail or composition comprising the remaining antibodies to glycophorin A, CD2, CD3, CD14, CD16, CD19, CD24, CD56, CD66b and IgE.
The present invention also relates to a kit useful for performing the processes of the invention comprising antibodies specific for glycophorin A, CD2, CD3, CD14, CD15, CD16, CD19, CD24, CD56, CD66b and IgE and instructions for performing the process of the invention.
Other features and advantages of the present invention will become apparent from the following detailed description.. It should be understood, however, that the detailed description and the specific examples while indicating preferred embodiments of the invention are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.